Neurochemical, molecular, immunohistochemical and behavioral methods were used to examine the in vivo effects of the neuropeptide galanin on central 5-HT neurotransmission and on 5-HT(1A) receptor-mediated responses. Intraventricularly infused galanin caused a long-lasting and dose-dependent reduction of basal extracellular 5-HT levels in the ventral hippocampus of awake rats as measured by microdialysis. Infusion of galanin into the dorsal raphe nucleus (DRN), but not intra-hippocampally, reduced 5-HT release. The effect of i.c.v. galanin on 5-HT release was blocked by the galanin receptor antagonist M35, acting most likely via galanin receptors at the level of the DRN. Galanin also reduced the levels of tryptophanhydroxylase mRNA in the DRN. Therefore, the effects of galanin on 5-HT(1A) receptor-mediated responses were further investigated. Surprisingly, galanin significantly attenuated the reduction of hippocampal 5-HT release induced by systemic injection of the 5-HT(1A) receptor agonist 8-OH-DPAT. Galanin also attenuated 8-OH-DPAT-induced hypothermia and locomotor activity in rats. These results indicate that galanin has important inhibitory actions on central 5-HT neurotransmission and on 5-HT(1A) receptor-mediated events.